Genetically engineered fungus
bites back at the crops it's meant to save

FOR the first time, a fungus has been genetically modified
to be more deadly to the weed it blights. The snag is that the
GM fungus kills crop plants as well. While the modified fungus
will not be released as a result of the findings, the case shows
how genetic modification can have unintended consequences. It
is also proof, were any needed, that biotechnology could be harnessed
to create weapons that attack crops (see page 4). The fungus was
modified to attack velvetleaf weed (Abutilon theophrasti). As
it is a close relative of cotton, most weedkillers that target
it destroy the crop as well as the weed. "Herbicides don't
work, and that's where you have to head in with biocontrol,"
says Jonathan Gressel, a plant physiologist at the Weizmann Institute
of Science in Rehovot, Israel. In theory, diseases are ideal for
biocontrol, because many infect just one plant species. The US,
for example, is testing funguses that target coca plants or opium
poppies. But diseases such as the fungus that causes anthracnose
in velvetleaf tend not to make good killers, as any that eradicates
its host is itself doomed. So Gressel's team decided to give the
anthracnose fungus (Colletotrichum coccodes) a killer punch by
adding a gene for a toxin from another fungus, Fusarium. The modified
fungus was indeed much more lethal to velvetleaf seedlings in
greenhouse experiments, they report in NatureBiotechnology (DOI:
10 .1038/nbt743). "This puts it over the brink to something
that would be useful," says Gressel. But the enhanced fungus
also killed off tomato and tobacco seedlings, neither of which
would normally be affected by the anthracnose fungus. This is
exactly the kind of unexpected consequence of a genetic modification
that opponents of GM have been warning about, although the case
also shows that such effects can be detected at an early stage.
"This business of putting in a toxin raises a red flag,"
says lane Rissler of the Union of Concerned Scientists in Washington
DC.

"This is exactly the kind of unexpected consequence that
opponents of genetic engineeringwarn about"

To allay these fears, Gressel suggests further "fail-safe"
modifications to any such fungus before it is tested outside a
sealed greenhouse. Removing the genes for sexual reproduction
would prevent it passing on any added virulence genes to related
fungi that attack other plants. And removing the genes for spore
formation would prevent it spreading via the air, and ensure it
died out completely each winter. Even these measures, however,
may not be foolproof For example, the fungus may survive without
spores, especially in moist tropical regions, says plant pathologist
Alan Watson at McGill University in Montreal. There may be other,
safer ways to boost the killing power of biocontrol agents. Watson
has patented a mixture of anthracnose fungi and low doses of ordinary
herbicides for weed control. The herbicide interferes with the
weed's normal defences against disease, allowing the fungus to
get the upper hand. It has yet to be tested on cotton fields,
however. Bob Holmes 0

Body gets a healing boost from
its inner electric fields

THE body's ability to heal itself partly depends on natural
electric fields, it has been proved for the first time. The work
could lead to the development of drugs that speed up healing by
enhancing these natural fields. Cell migration and division plays
a key role in development and healing. Most research in this area
is on chemical factors, but several studies have shown that applying
electric fields can affect migration and division as well. After
successful trials in animals, for example, a team in the US is
trying to encourage the healing of spinal cord injuries by applying
an extemal field. To discover whether the body's natural electric
fields play a role in healing, Colin McCaig's team at the University
of Aberdeen in Scotland looked at rat corneas. In an undamaged
cornea, cells pump positively charged ions into the comea and
push negatively charged ions out, creating an electrical potential
Of 40 millivolts. But in damaged areas this voltage disappears,
setting up an electric field between damaged and intact areas
that stretches for half a millimetre along the surface of the
injured cornea.

The researchers used a variety of chemicals to nullify or enhance
this natural field. Dividing cells tended to line up at right
angles to the field. But when the fields were nullified, the cells
aligned randomly, they report in Proceedings of the National Academy
ofsciences (DOI:10.1073/ pnaS202235299). McCaig says that the
electric field may attract positively charged proteins or lipids
in the membranes of cells. What's more, the team found that healing
was faster when the field was boosted, and slower when it was
decreased. Stronger fields also encourage cells to divide, though
how they do this isn't clear.

McCaig says the work is the first demonstration that the body's
natural electric fields play a key role in healing. "It's
a big step forward to help biologists believe that the fields
are important.' It's possible that drugs could be developed that
enhance the natural field around wounds, boosting healing. This
may prove more effective than applying an external field that
doesn't have the same shape as a natural field. Jon Copley and
Michael Le Page

Saving up Mutations

Evolution is a slow, painstaking process. But have plants and
animals found a way of seizing the throttle to get them out of
a tight spot? BobHolmes reports

DESPITE its universal role in biology, evolution still poses
some pretty perplexing questions. Take changes in body form. Every
tree or beetle or mouse looks the way it does because thousands
of genes turned on at exactly the right time and place to guide
the organism from single cell to adulthood. But if body plans
are the product of such intricately orchestrated programs, how
can evolution ever conjure up new ones? Any slight perturbation
would surely send a species tumbling from its evolutionary peak
into the barren valleys beneath. Plants and animals may have hit
on an ingenious solution - bottling up evolution for times when
they really need it. By squirrelling away genetic mutations, the
raw material of evolution, and releasing them all at once, species
may be able to leap from peak to evolutionary peak without ever
having to slog through the valleys between. This happy knack increases
their odds of surviving stressful conditions - nothing less than
evolution on demand. On the face of it, the idea sounds like biological
heresy. Plants and animals couldn't have that sort of control
over the random process underlying evolution, could they? Surprisingly,
they could. Over the past few years, a handful of lab experiments
have thrown up convincing evidence that organisms really can save
up mutations for a rainy day. If the same thing happens in nature,
then plants and animals have hit on a way to 9-9 seize the throttle
of evolution, accelerating it when necessary and slowing it down
when not. Their storehouse of mutations may also prove to be a
treasure trove of new genes for drug hunters to plunder or, equally,
the time-bomb that helps explain the diseases of old age. The
lead actor in this iconoclastic drama is a so-called "chaperone"
protein called hspgo. one of the most abundant proteins in animals,
plants and fungi, hspgo's job is to bind to unstable proteins
and help them maintain their correct shape. In this role, hspgo
is rather like a valet, tidying up proteins that would otherwise
become dishevelled by environmental insults such as high temperatures.
Hence the chaperones'other name, "heat shock proteins".
But hspgo is also a crucial regulator of development. As a way
of silencing proteins until their services are required, cells
deliberately make some proteins unstable, especially certain ones
that regulate developmental pathways. One of hspgo's jobs is to
hold some of these proteins in the "standby" position.
"it works on just about every [developmental] pathway you
can imagine," says Susan Lindquist, director of the Whitehead
Institute for Biomedical Research in Cambridge, Massachusetts.
The first hint that hspgo's job gives it unusual leverage over
evolution came four years ago, when Lindquist and her team mate
Suzanne Rutherford were working at the University of Chicago.
They noticed that fruit flies carrying a mutant copy of the hspgo
gene sometimes had offspring that looked very weird indeed. "We
had eyes that grew out from the head in a stalk-like pattem, we
had bristles in the wrong places, wings with different venation
pattems and shapes, abdomens that were partly folded over, legs
that were different shapes - virtually every structure in the
adult fly was affected," says Lindquist. The same abnormalities
showed up in normal flies doped in the larval stage with geldanamycin,
a drug that [email protected]@th the action of hspgo (Nature, vOl 396,
P 336). That wasn't too surprising, given hspgo's pivotal role
in development. But when the researchers looked more closely at
their abnormal flies, they saw something much more interesting:
each set of parents tended to produce offspring with a distinctiv6
set of abnormalities, different from those in unrelated fly lineages.
One might have deformed legs, another strangely positioned eyes
and stunted wings. If the flies'bizarre body plans were simply
down to a shortage of hspgo playing havoc with their developmental
pathways, then the defects should have been scattered randomly
throughout the whole fly population. But they weren't. There was
something else going on, and the researchers thought they knew
what it was. They suggested that the familial abnormalities were
caused by "cryptic" genetic defects that had lain hidden
for generations and only showed up when hspgo stopped doing its
job. That would explain Why each lineage sported a different set
of abnormalities: each one had its own, unique collection of hidden
defects.

This makes sense given hspgo's way of working. When all is
well, the researchers proposed, hspgo goes about its usual business
of keeping unstable proteins in working order. it does this so
efficiently that it can even tuck into shape proteins with minor
mutations that would otherwise alter their shape and, therefore,
their functions. In effect, hspgo papers over the flaws in an
organism's genome by keeping mutations hidden harmlessly away
- including mutations in the genes that regulate development.
Over the generations, a lineage of flies can accumulate many minor
mutations that never see the light of day. in times of stress,
though, this patina of orderliness breaks down. High temperatures,
noxious chemicals, and a host of other stresses can cause an epidemic
of protein misfolding. Faced with a dramatically increased workload,
hspgo can no longer keep up and, as a result, malformed proteins
go unrepaired. That means stress can lead to the sudden unmasking
of hidden mutations. If it happens during larval development or
metamorphosis, hidden mutations in developmental genes can produce
abrupt changes in shape and form. Remarkably, then, hspgo acts
as both a capacitor for storing genetic variation and the trigger
that releases it. It therefore gives evolutionary biologists a
convincing molecular explanation for evolutionary change. "This
looks like something that's going to put evolutionary theory on
a firmer ground in terms of mechanism," says Massimo Pigliucci,
an evolutionary ecologist at the University of Tennessee in Knoxville.
Earlier this year, Lindquist's team announced that hspgo performs
the same trick in the thale cress, Arabidopsis thaliana - making
it likely that many other organisms store variation in this way,
too.

"This is almost too good to be true. Justwhen you need
variation - the raw material of evolution - its there?"

Once again, drugs that interfere with hspgo caused some seedlings
to develop abnormally. "We had roots that grew up instead
of down, changes in the number of root hairs, leaves that developed
almost like pine needles, leaves that curled up or down, leaves
that became pigmented - all sorts of things," says Lindquist.
Many of the variants looked as if they might be helpful to plants
trying to adapt to new environments - more or fewer root hairs,
for example, might be appropriate for different soil types and
moisture conditions - though the researchers did not test this
directly. Again, different inbred lines of the plants produced
seedlings with different sets of abnormalities, and they showed
the same characteristic abnormalities when grown at higher temperatures,
even without the drug (Nature, vel 417, p 6i8). Hspgo seemed to
be concealing cryptic mutations in plants, too. What's more, once
these new variants come out of the woodwork, the useful ones are
likely to stick around even after the environmental stress has
disappeared. When Lindquist'i team bred fruit flies while selecting
for certain abnormalities, after just a few generations the flies
hung onto their new shapes even when hspgo was at full function
again. Lindquist believes that selecting for these traits, which
are the result of many genes working together, prompts the selected
lines to accumulate more and more of the desirable gene variants,
until eventually the flies exceed a threshold where the trait
becomes independent of hspgo. That's important, because it means
organisms won't lose useful mutations once the stress evaporates.
The upshot of this is that species seem to have a mechanism for
delivering variation - the raw material of evolution - just at
the time they need to adapt to a changing environment. "It's
almost too good to be true. just when you need variation, it's
there," says Charles Knight, an evolutionary physiologist
at the Max Planck Institute for Chemical Ecology in jena, Germany.
Maybe so, but Lindquist's results actually sit quite snugly with
existing theories of evolution and development. As early as the
1940s, British biologist C. H. Waddington suggested that organisms
must have ways of buffering mutations that could disrupt their
development. Though Waddington had some experimental evidence,
many biologists remained sceptical, because no one knew how such
buffering could arise. Lindquist's work provides the first molecular
explanation. "The hspgo work came as a real surprise,"
says Brian Hall, an evolutionary developmental biologist at Dalhousie
University ih Halifax, Nova Scotia. "Here's this molecule
we've known about for quite some time that could play this really
fascinating role." Hspgo's ability to store and release mutations
also helps resolve a long-standing evolutionary puzzle - how species
can make the transition from one body plan to another when intermediate
forms would seem to be dangerously maladapted. By storing genetic
variation and releasing it all at once, a species may be able
to muster the raw material for big evolutionary leaps. These larger
leaps could increase a species'chances of finding a design that's
better adapted to its new conditions, says Lindquist. It may even
have been one of the driving forces behind some of the bursts
of rapid diversification found in the fossil record - the so-called
"punctuated equilibrium" model of evolution popularised
by the late palaeontologist Stephen Jay Gould. For all the advantages
of such a system, though, Lindquist has backed away from the suggestion
- which she and Rutherford hinted at in their first paper - that
this storage-and- release mechanism might have been shaped for
that purpose by natural selection.

"This looks like somethingthat's going to put evolutionary
theory on firmer ground in terms of mechanism"

"We're not by any means saying that it evolved for the
sake of evolvability," she says now. Most other researchers
agree. "It's actually very difficult to think up cases in
which systems evolve in order to make evolution more efficient,"
says Nicholas Barton, an evolutionary geneticist at the University
of Edinburgh. "It's not impossible that that sort of thing
can happen, but it takes careful argument to justify it.' Instead,
hspgo's buffering ability most likely arose as an incidental by-product
of its main role in protecting proteins against environmental
stress. But buffering you from genetic mutation may in the end
have a less desirable side effect too: one researcher suggests
that overloading your chaperone system as you get older may be
one cause of the diseases of ageing (see "The shock of the
old"). While Lindquist's experiments have focused on hspgo,
it is unlikely to be the only protein playing this evolutionary
game. "I think this is the tip of the iceberg. There are
going to be many things that buffer genetic variation," says
Lindquist. Hspgo is just one of a whole platoon of heat shock
proteins, and other molecules may also act in similar ways. Earlier
this year, for example, researchers at the University of Valencia
reported that an hsp dubbed GroEL can repress harmful mutations
in the bacterium E. coli (Nature, vol 4i7, P 398). Though even
sceptics say her experiments are impeccable and the buffering
mechanism she describes is fascinating, Lindquist herself wouldn't
claim organisms necessarily enjoy these payoffs in the real world.
"For actual long-term evolution to occur, first of all one
of the phenotypes that's uncovered needs to be beneficial,"
says University of Chicago biologist Martin Feder. Then it has
got to hang around long enough to shed its dependency on hsp and
become abundant in the population. That means the organism expressing
it must find a mate whose genes allow this trait to appear. "While
not impossible, these events are fairly improbable," Feder
concludes. Then, too, fruit flies and thale cress are hardly typical
of most species in the wild. Besides their many generations of
adaptation to life in the lab, these two species became geneticists'favourites
partly because of their unusually short life cycles, which means
they accumulate mutations more quickly than other plants or animals.
More sedate species might never gather enough hidden mutations
for this mechanism to be important. "It's possible that the
role of heat shock proteins may be overestimated in organisms
that reproduce that fast," says Pigliucci. Nor is it clear
that providing more variation will actually prompt organisms to
evolve faster. Most natural populations already express ample
genetic variation to support evolution, argues Barton: "Even
if heritable variation were much lower than it really is, we would
be able to account for evolution. You don't have to suppose that
organisms are sitting around waiting for variability to come up.'
As evidence, he notes that if you select organisms for almost
any trait you choose, they'll respond - and faster than you usually
see evolution proceed in the fossil record. These will remain
open questions until someone can use the hsp system to produce
useful adaptations. Feder says he and Lindquist have talked about
trying this critical experiment on yeast, but their plans got
pushed aside during Lindquist's recent move from Chicago to Massachusetts.
No matter how important hspgo's masking of mutations turns out
to be in real-world evolution, Lindquist's experiments seem certain
to provide other scientists with a valuable tool. Plant breeders,
for example, may be able to expose the hidden variation within
a crop species as an altemative to costly and time-consuming techniques
such as genetic engineering and cross-breeding with wild relatives.
Developmental geneticists may be able to use a similar approach
to pick apart the evolution of development. Hall, for example,
plans to see whether he can uncover hidden variation in a vertebrate,
the dwarf African frog. If so, he hopes to use drugs to block
hspgo at different stages of the frog's life cycle to gauge the
variation at each stage. Such a snapshot, he thinks, would reveal
which developmental stages allow the most leeway for innovation
and which are most conservative. All in all, Lindquist's results
have evolutionary biologists buzzing. "This could open the
floodgates to a lot of follow-up," says Pigliucci. Not a
bad yield from a bunch of deformed fruit flies.